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. 2023 Jan 20;40(5):1163–1171. doi: 10.1007/s10815-023-02727-w

Autologous bone marrow-derived nucleated cell (aBMNC) transplantation improves endometrial function in patients with refractory Asherman’s syndrome or with thin and dysfunctional endometrium

Gurkan Arikan 1,✉, Volkan Turan 1,2, Meryem Kurekeken 1,3, Hasan Sami Goksoy 4, Zeynep Dogusan 5
PMCID: PMC10239402  PMID: 36662355

Abstract

Purpose

The purpose was to evaluate the effect of intrauterine injection of aBMNC on the endometrial function in patients with refractory Asherman’s syndrome (AS) and/or thin and dysfunctional endometrium (TE).

Study design

This is a prospective, experimental, non-controlled study

Material and methods

The study was carried out between December 2018 and December 2020 on 20 patients, who were of age < 45 years and had oligo/amenorrhea and primary infertility due to refractory AS and/or TE. One hundred ml BM was extracted. aBMNC cells were separated according to generic volume reduction protocol by using the Cell Separation System SEPAX S-100 table top centrifuge system. We have evaluated CD34+, mononuclear cell (MNC), and total nucleated cell (TNC) counts. The transplantation aBMNC was performed by two intrauterine injections at an interval of one week, transvaginally into the endometrial–myometrial junction by an ovum aspiration needle. Midcyclic endometrial thickness (ET) and gestations after transplantation were evaluated.

Results

The mean TNC, MNC, and CD34+ cells were 11.55 ± 4.7 × 108, 3.85 ± 2.01 × 108, and 7.00 ± 2.88 × 106 at first injection, respectively, and 6.85 ± 2.67 × 108, 2.04 ± 1.11 × 108, and 3.44 ± 1.31 × 106 at second injection, respectively.

The maximum posttransplantation ET was significantly higher than the maximum pretransplantation ET: 2.97 ± 0.48 vs. 5.76 ± 1.19 (mean ± standard deviation, p < 0.01). Twelve patients had frozen-thaw embryo transfers after the study.

In 42% (n = 5 of 12) of the patients, pregnancy was achieved. One of the five patients delivered a healthy baby at term.

Conclusions

Autologous BMNC transplantation may contribute to endometrial function in patients with AS and/or TE.

Keywords: Autologous bone marrow, Total nucleated cells, Stem cells, Endometrium, Asherman’s syndrome, Thin endometrium

Introduction

Asherman’s syndrome (AS) may be caused by the destruction of the endometrium due to repeated or aggressive uterine cavity surgeries [1–3], which leads to fibrotic adhesions and loss of glands, followed by partial or complete loss of functional endometrium in the cavity. The described pathology is usually aseptic and is based on the surgical damage of the endometrium [2]. Its prevalance varies from 2 to 22% in infertile women [3]. The patients suffer under hypomenorrhea, amenorrhea, infertility, recurrent pregnancy loss, or abnormal placentation. Minimal or mild adhesions are easily divided and usually do not affect embryo implantation. Moreover, in severe AS, the tubes may be blocked and the loss of functional endometrium is detrimental to the implantation of the embryo.

Similarly, thin and dysfunctional endometrium (TE) or endometrial atrophy (EA) has been described as another rare condition in which the endometrium remains refractory to estrogen treatment. The frequency is about 0.5 % in patients with infertility [4, 5]. TE may be idiopatic or due to uterine surgeries, ovarian stimulation with clomiphen citrate, prolonged use of progesteron, or combined oral contraceptive pills [5].

Several medical and surgical therapy methods have been introduced to treat the latter, such as prolonged estrogen administration, vaginal sildenafil, vitamin E, pentoxyfylline, GnRH-agonist supplementation [4], endometrial curretage, hysteroscopic adhesiolysis, and anti-adhesive barriers, yet none have proved effective.

Endometrial stem/progenitor cells reside in the basalis layer of the endometrium and are responsible for regenerating endometrium in each cyle. Their diminished number or function may result in a thin dysregulated endometrium, leading to embryo implantation failure, as in AS and TE [6].

Stem cells have effects on tissue repair by homing in on the injured site, recruiting other cells by secreting chemokines, modulating the immune system, differentiating into other types of cells, proliferating into daughter cells, and potentially having antimicrobial activity. Circulating bone marrow-derived hemopoietic stem cells, mesenchymal stem cells, and endothelial progenitor cells integrate into damaged tissues and transdifferentiate into host tissues, including endometrium [7, 8].

Bone marrow-derived cells consist of a heterogeneous mix of mesenchymal stem cells (MSC), hematopoietic progenitor cells (HPC), endothelial progenitor cells (EPC), immature monocytes, and lymphocytes [9, 10]. Circulating BM-derived CD34+ CD45+ cell release angiogenic and chemo-attractant factors and are powerful angiogenic agents [11]. Once they migrate into ischemic tissues, they recruit immune cells (neutrophils and monocytes) and activate other cells that participate in the inflammatory response and contribute to vascular remodeling [12]. Furthermore BM-immune precursor cells are thought to have an active role in angiogenesis and/or arteriogenesis itself [13]. For example, neutrophils do not only participate in inflammation but can also promote vascularization by inducing angiogenesis via a proangiogenic phenotype [12, 14, 15].

Administration of bone marrow-derived stem cells (BMSC) into tissues and organs has been shown to contribute to the repair and regeneration, in animal [16–23] and also in human endometrium [24–28]. There is consensus on the regenerative effect of bone marrow-derived stem cell-based therapy on endometrium. However, the existing literature contains little data on small groups of patients [24–31].

Based on previous data, we conducted a study designed to evaluate the safety, feasibility, and efficacy of transvaginal intrauterine injection of aBMNCs in patients with refractory AS or TE, in terms of sonografic and functional improvement of the endometrium in ART cycles.

Material and methods

Selection of participants

This study was approved by the institutional Ethics Committee (TRN 047, 26.12.2018). The procedures used in the study were in accordance with guidelines of the Helsinki Declaration of 2000 on human experimentation. All participants provided written informed consent.

The study was carried out between December 2018 and December 2020 in a tertiary care university clinic with the cooperation of the departments of obstetrics and gynecology, haematology, and stem cell laboratory.

Patients of age < 45 years with oligo/amenorrhea and primary infertility due to refractory Asherman’s syndrome and/or TE (endometrial thickness < 5 mm), for whom standard treatment options had failed, were offered to participate. All participants had undergone a hysteroscopy within 8 weeks prior to enrollment into the study.

Exclusion criteria were chronic hematological diseases, hydrosalpinx, persisting uterine fluid, and chronic genital infections such as tuberculosis.

Autologous bone marrow (aBM) extraction and cell analysis

One hundred milliliter of the bone marrow was harvested by puncture of sacrum under local anesthesia and autologous bone marrow nucleated cells (aBMNC) were separated according to generic volume reduction protocol by using the Cell Separation System SEPAX 2 S-100 table top centrifuge system (Biosafe Group SA, Eysins, Switzerland). The SEPAX 2 S-100 system allows the automated processing of cell components in a functionally closed and sterile environment. The SEPAX 2 S-100 provides centrifugal and axial displacement drive to the chamber on the single use separation kit, as well as drive to the directional valves. The cell separation process is permanently monitored by an optical sensor, fully automated, and completed within 15 to 20 minutes. The generic volume reduction protocol uses a single sedimentation step with centrifugal force of 960 × g and concentrates the final cell product. The nucleated cells are separated reliably and cost-effectively.

The final 15 ml concentrated aBM product was sampled in a 5-ml standard vacutainer tube containing EDTA. Quantitative assessment of cell population was performed by Celltac α cell counter (Nihon Kohden, Japan). A stem kit from the Beckman Coulter Navios (Miami, FL, USA) was used for the cell labelling with CD34-PE, CD45-FITC, 7-AAD, and Stem Count fluorospheres (Stem-Kit Reagents, IM3630). The analysis protocol was used for detection of CD34+ CD45+ (progenitor) cell counts and cell viability to define the aBM product. We also analyzed percentages of the granulocytic, monocytic, and lymphocytic compartments of aBMNCs by flow cytometer according to the manufacturer’s protocol (Beckman Coulter, France). We incubated 50 microliters of BMNC sample with fluorescence-labeled monoclonal antibodies against CD45 (FITC, A07782), CD14 (PE, A07764), CD19 (ECD, A07770), CD3 (PC5, A07749), and CD16 (PC7, 6607118).

Half of the harvested aBMNCs was prepared for the first transvaginal intrauterine injection. The rest of the aBMNC was cryopreserved in a final concentration of 10% dimethyl sulfoxide (DMSO) and autologous plasma by a controlled rate freezer. After the cryopreservation procedure, the aBMNCs were stored in the vapor phase over liquid nitrogen, where the temperature was constantly below – 150 °C.

After one week, aBMNCs were thawed in a water bath at 37 °C for 1–3 min and were sent to the IVF center for the second injection of aBMNCs. Postthaw viable CD34+ CD45+ cell counts were determined by flow cytometry.

Transplantation of aBMNCs

The transplantation protocol consisted of two injections of aBMNCs, one between the 8th and 10th days of the menstrual cycle and the other between the 15th and 17th days of the menstrual cycle. The first half of the harvested aBMNCs was injected transvaginally into the junctional zone of the endometrial–myometrial interface by an ovum aspiration needle (Cook Nr. 17) under sonografic guidance and general anesthesia. A maximum volume of 35 ml was delivered at 5–8 sites around the uterine cavity. The second half of the aBMNCs was cryopreserved and transplanted one week after the first procedure, also as described above.

Follow-up

All patients were given hormone replacement therapy (HRT) (6 mg estradiol daily, Estrofem 2 mg Tablets, Novo Nordisk Pharma GmbH, Germany) before the transplantation of the aBM pruduct. HRT was given 6 weeks after the last surgical procedure or until the next menstrual bleeding.

Midcyclic endometrial thickness (ET) was assessed by vaginal ultrasound once in each three intervals (between the 2nd and the 4th month, between the 5th and the 7th month, and between the 8th and the 10th month) after the aBMNC transplantation and during the ART cycles.

Information on ART cycles and gestations was collected.

Statistical analysis

Statistical analysis was performed using SPSS 22.0 Software (IBM, MD, USA).

Student’s t test with paired data was used for comparison between two groups of normally distributed parameters and Mann–Whitney U test was used for comparison between two groups of parameters that did not show normal distribution. Fisher’s exact test was used to compare qualitative data. A p value obtained in a two-tailed test ≤ 0.05was considered statistically significant.

Results

Twenty patients aged < 45 years with infertility and refractory AS or TE were enrolled into the study between December 2018 and December 2020.

The patients’ characteristics are shown in Table 1. All had a history of hysteroscopic adhesiolysis and in vitro fertilization therapies and the endometrium remained refractory (< 5 mm) under HRT. The number of embryo transfers before the study was 3; (1–4) (median; (range)). All patients were planned either for frozen thaw embryo transfer or for new ART trials following our study. In all patients severe AS and or EA were diagnosed by hysteroscopy. A hysterosalpingography prior to our study showed synechiae (filling defects) or irregular endometrial contours in 12 (60%) patients (Fig. 1a and b) and bilateral tubal blockage in 15 (75 %) patients. The tubal blockage was considered to be a result of occlusion of ostia in severe AS, as previously described [32] or of an additional tubal disease.

Table 1.

Baseline characteristics of the patients; mean ± SD (%).

Baseline characteristics
Age (mean ± SD) 36.6 ± 5.59
BMI (kg/m2) 25.66 ± 2.56
Number of previous hysteroscopies (mean ± SD) 3.33 ± 1.6
Number of previous fresh or frozen-thaw embryo transfers (mean ± SD) 2.9 ± 0.67
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BMI, body mass index; SD, standard deviation

Fig. 1.

Fig. 1

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Hysterosalpingography of patient #2 (a) and patient #6 (b) showing synechiae, irregular endometrial contours, and occluded ostia

Table 2 shows the mean count of TNC, MNCs, the CD34+CD45+ cells in the first and second injections, and their viability, respectively. The mean count of MNC in the first and in the second injection was 3.85 ± 2.01 × 108 and 2.04 ± 1.11 × 108, respectively. The mean count of CD34+ cells transplanted in the first and in the second injection was 7.00 ± 2.88 × 106 and 3.44 ± 1.31 × 106, respectively, and the mean viability was roughly 98% and 54%, respectively.

Table 2.

. The mean count of transplanted total nucleated cells (TNC), mononuclear cells (MNC), CD34+ cells in the first and second aBMNC injections, and their viability (%; mean ± standard deviation), respectively

First injection Second injection
TNC 11.55 ± 4.7 × 108 6.85 ± 2.67 × 108
MNC 3.85 ± 2.01 × 108 2.04 ± 1.11 × 108
CD34+ cells 7.00 ± 2.88 × 106 3.44 ± 1.31 × 106
Viability of CD34+ cells 98.3 ± 1.58% 54.5 ± 16.0%
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The mean percentages of CD3+ cells (T lymphocytes), CD19+ (B lymphocytes), CD14+ cells (monocytes), and CD16+ cells (granulocytes) were 74,95 ± 7,54, 12,8 ± 7,33, 87,0 ± 4,37 %, 72,8 ± 8,18, respectively.

The maximum posttransplantation ET was significantly higher than the maximum pretransplantation ET: 2.97 ± 0.48 vs. 5.76 ± 1.19 (mean ± standard deviation, p < 0.01). Figure 2 shows the maximum ET of the patients in pre- and posttransplantation period. An improvement of maximum ET was observed in all but patient #7 after the aBMNC transplantation. In 50% of the patients, we observed an uneven improvement of endometrium thickness, possibly due to remaining adhesions. In others the improvement of ET was uniform and consistent (Fig. 3a–c).

Fig. 2.

Fig. 2

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Maximum endometrial thickness of the patients pre- and posttransplantation (mm)

Fig. 3.

Fig. 3

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(a) Ultrasound image of a patient pretransplantation with maximum ET of 3 mm. (b) Uneven improvement of endometrium 3 months posttransplantation, with maximum ET of 5.8 mm. (c) Uniform and consistent improvement of the endometrium in another patient 6 months posttransplantation

Twelve patients had frozen-thaw embryo transfers after the study. Eight patients had blastocyst transfers, and 4 had embryos transferred in cleavage status. In 42% (n = 5 of 12) of the patients, pregnancy was possible to achieve after ART. One of the patients delivered a healthy baby at term. Three patients had biochemical pregnancy, and one had a missed abortion. All pregnancies were achieved after embryo transfers in the stage of blastocyst.

Four patients dropped out during follow-up. Four out of the remaining 16 patients were initially amenorrheic and 12 had scant menstrual bleeding. After the aBMNC transplantations, we observed that the menstrual cycles were restored in 3 of the 4 amenorrheic women. Ten out of 12 women with scant menstrual bleeding reported an increase in the flow of menstrual bleeding. The further details of menstrual flow were not documented, since this was not defined as an outcome criterion.

The patients’ age, body mass index, TNC, MNC, CD34+ cell counts, and viability rate of the cells had no statistically significant impact on the outcome criteria (p > 0.05). No adverse effects were observed.

Discussion

This preliminary prospective longitudinal study showed that the transvaginal intrauterine injection of aBMNCs improves endometrial function in patients with refractory Asherman’s syndrome and/or with thin and dysfunctional endometrium.

The technique of extracting stem cells directly from bone marrow and their intrauterine transplantation are feasible and safe procedures.

The maximum posttransplantation ET was significantly higher than the maximum pretransplantation ET: 2.97 ± 0.48 vs. 5.76 ± 1.19 (mean ± standard deviation, p < 0.01). A pregnancy was achieved after ART in 42 % of the patients. One patient delivered a healthy baby at term. 3 patients had biochemical pregnancies and one had a missed abortion.

The first study to report bone marrow stem cells (BMSC) playing a role in the regeneration of the endometrium was published in 2004 [24]. The authors detected donor BMSC in the endometrium of patient, who had received HLA-mismatched bone marrow transplants. In 2011, Nagori et al. described the first successful BMSC therapy of a patient with AS and infertility [25]. The authors extracted bone marrow and instilled endometrial angiogenic cells transcervically into the uterus. They reported an increase in ET from 3 to 7.1 mm and a conception after subsequent ART with oocyte donation. Singh et al. [28] evaluated the effects of BM-MNC implantation in patients with refractory AS. CD34+ BM-MNCs were injected into subendometrial zone and observed the improvement of endometrial thickness from 1.38 ± 0.39 to 5.46 ± 1.4 mm.

A larger experimental, non-controlled study was carried out [27], in which 18 patients with refractory AS or endometrial atrophy (EA) underwent autologous BMSC therapy. The study plan consisted of bone marrow stem cell mobilization by granulocyte colony stimulation factor injection, isolation of CD133+ cells by peripheral blood aphaeresis, stem cell administration into the spiral arterioles by catheterization of femoral artery, and an adhesion scoring by hysteroscopy after the trial. AS and EA patients showed a significant increase of endometrial thickness two months after stemcell therapy (from 4.3 to 6.7 mm; p < 0.004 and from 4.2 to 5.7 mm; p < 0.03, respectively). Seven pregnancies were obtained after 14 embryo transfers, resulting in 1 baby born. Three patients became pregnant spontaneously, resulting in one baby boy born, one ongoing pregnancy, and a miscarriage at the time of publication. The authors reported spontaneous pregnancies after stem cell therapies [27–29], indicating that some patients with mild AS and patent tubes were enrolled. Singh et al. [28, 29] have also recruited patients with adhesions after or during genital tuberculosis. In our study, all patients had posttraumatic and severe AS and all needed ART for conception. Therefore, the endometrial disease of patients in the previous studies may not be considered to be comparable to ours. However, in our study, the improvement of endometrial thickness was similar to the previous studies and the rate of pregnancy after ART was comparable [27–29]. The transvaginal intrauterine injection of aBMNCs in our study was less invasive and less costly, since no peripheral blood apheresis, cell sorter CliniMACS and radiological interventions were needed. Nevertheless, the closed system of SEPAX provided a sterile high-quality isolation of nucleated bone marrow cells.

Various stem cell markers have been tried in previously published studies, but currently, there is no consensus on which stem cell marker is most suitable for endometrial regeneration. CD9, CD44, and CD90 were used as surface marker of angiogenic stem cells [25]. CD133+ is a proven endothelial progenitor cell marker, which was used to isolate angiogenic stem cells by Santa Maria et al. [27]. CD34+ is a surface of hematopoietic and also endothelial progenitor cells, which helps in angiogenesis and tissue repair [33, 34]. CD34+ cells have the potential for neovascularization in ischemic tissue to restore microcirculation and improve tissue perfusion [35]. Singh et al. [28] quantified the CD34 marker only; however, they transplanted the whole BM-MNC product and found no correlation between the cell counts and clinical outcome and refrained from evaluating the proportion of CD34+ cells in BM-MNC in a later study [29]. In contrast, others found that the number of applied CD34+ stem cells was an independent predictor of response to BM-MNCs in a controlled study in patients with refractory diabetic sensorimotor polyneuropathy [36].

The authors used different techniques to isolate BMSC and the volume of bone marrow extraction and cell counts vary highly among published studies. Santa Maria et al. isolated a minimum of 50 million CD133+ cells for systemic administration by peripheral blood apheresis and isolation of the cells by using the cell sorter CliniMACS. In some patients, it was necessary to process two to three times the blood volume (each 10 liters). Singh et al. used the Ficoll density separation method and reported to have aspirated 30 ml of BM, gained mean 103.3 ± 20.45 × 106 MNCs; and the mean count of CD34+ cells were 58,800 (0.20 % of MNCs). We have aspirated 100 ml of BM, and the median MNC and CD34+ cell counts were considerably higher: 3.28 ± 2 × 108 and 7.44 ± 2.88 × 106 (2.26% of MNCs), respectively. Similar to others (28), we found no correlation between cell counts and clinical outcome. The documentation of the counts of TNC, MNC, and CD34+ cells will help the evaluation of correlations to clinical data in the future.

Ours is the first study on patients with AS and TE, where all nucleated cells were isolated by the SEPAX system and transplanted for endometrial regeneration. The subpopulations of nucleated cells accompanying the stem cells may have contributed to regeneration processes [11–15]; bone marrow-derived cells consist of a heterogeneous mix of MSC, HPC, EPC, immature monocytes, and lymphocytes [9, 10]. In addition to angiogenic and chemo-attractant factors released by CD34+ cells [11], these cell populations recruit immune cells (neutrophils and monocytes) and activate other cells that participate in the inflammatory response and contribute to vascular remodeling [12]. It has been shown that the conditioned medium, which contains the secreted factors collected from T helper 2 and T helper 17 Tcell phenotypes, enhances angiogenesis in vitro and in vivo [37]. Furthermore, macrophages secrete pro-angiogenic factors such as vascular endothelial growth factor (VEGF) and fibroblast growth factor (FGF) leading to formation of new blood vessels [38].

The beneficial effects of direct administration of stem cells may also result partly from transdifferentiation of the BMSC and/or from injury-induced stimulation of poorly functioning endogenous endometrial stem/progenitor cells [6, 26]. The observations that the endometrial injury or serial biopsies preceding IVF-ET cycles may increase the pregnancy rate suggest that endometrial injury provokes an inflammatory and/or wound healing response with associated release of growth promoting cytokines inducing decidualization [39, 40]. Cervelló et al. have investigated the engraftment and proliferation of human CD133+ BM-MSC in mice [17] and observed a significant increase in cell proliferation and paracrine factors in the treated damaged horns relative to the untreated damaged horns, regardless of administration modality. Santamaria et al. reported that the positive effect of cell therapy on mensturation decreased 6 months after systemically administered stem therapy in patients with AS. In contrast, Singh et al. [29] observed that the improvement of endometrial thickness remained stable over 5 years in their longitudinal study after subendometrial administration of MNCs. Whether paracrine changes and tissue injury have beneficial effects compared to systematical administration route remains to be investigated in future studies.

This is the first study with repeated cell therapy in patients with endometrial disease. The poor engraftment of the transplanted cells is a universal phenomenon that has been observed with all types of cells studied [41–44]. Repeated cell administrations have cumulative beneficial effects and, as a result, are markedly more effective than a single administration [41]. However, all clinical studies in patients with AS performed heretofore have used one cell administration. There are several reasons for this insistence on single-dose treatments. First, it was thought that adequate regeneration could be attained simply by administering one (sufficiently large) dose of cells. However, as mentioned above, there is possibly minimal long-term engraftment, regardless of the cell type used. Secondly, the use of repeated cell injections is hindered by logistic, safety, financial, and regulatory issues. In our study setting, we did not have the limiting issues mentioned above. Second injection of aBMNCs was an additional but minimal intervention to our patients; we believe that this strategy increased the engraftment and enhanced the beneficial paracrine mechanisms.

Finally, dimethyl sulfoxide (DMSO) used in cryopreservation protocol was reported [45] to act through different unspecific changes, increase chemotactic response of hematopoietic cells, and contribute to homing and survival of stem cells in an animal model. The authors stated that a 10% DMSO could be considered a way of increasing homing and engraftement efficacy of transplanted stem cells and may be a viable alternative to already suggested strategies. The DMSO might have increased the regenerative effect of BMNCs at the second injection in our study.

Limitations of our study were small sample size, absence of a control group, short follow-up period, and drop-outs from the study due to exhausted fertility desire on the part of the patients or due to diminished ovarian reserves. Future comparative studies in younger patients with adequate ovarian reserves and preimplantation genetical testing are needed. Other sources of stem cells could be investigated to regenerate endometrium in patients with AS or TE, such as allogeneic endometrial stem/progenitor cells, autologous endometrial cells derived from induced pluripotent stem (iPS) cells, or umbilical cord-derived mesenchymal stem cells [46].

Conclusions

We conclude that the transplantation of autologous bone marrow nuclear cells by transvaginal intrauterine injection is a safe and feasible technique and improves the endometrium in patients with refractory Asherman’s syndrome and/or with thin and dysfunctional endometrium.

Acknowledgments

We thank Professor Erdal Karaoz (PhD) for his comments on previous and final versions of the manuscript and Ayşenur Kocaoglu and Beril Mensure Karakan, for their technical asistance.

Author contribution

All authors contributed to the study conception and design. The study was conducted by Gurkan Arikan. The transplantations of aBMNC were performed by Gurkan Arikan, Volkan Turan, and Meryem Kurekeken. The BM aspirations were performed by Hasan Sami Göksoy. The Cell laboratory was run by Zeynep Dogusan. Statistical analyses was performed by Volkan Turan and Meryem Kurekeken. All authors made substantial contributions to the conception and design of the work or to the acquisition, analysis, and interpretation of the data. The first draft of the manuscript was written by Gurkan Arikan and Volkan Turan, and all authors commented on previous versions of the manuscript. All authors have read and approved the final manuscript.

Funding

Partial financial support (for equipment and supplies only) was received from Yeniyüzyil University, Gaziosmanpasa Hospital.

Declarations

Ethics approval, consent to participate, and consent to publish

All procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional ethics committee and with the 1964 Helsinki Declaration and its later amendments or comparable ethical standards. The study was approved by the Bioethics Committee of the Medical University of Yeniyüzyil University (TRN 047, 26.12.2018). Informed consents for participation and for publication of the data were obtained from all patients.

Competing interests

The authors declare no competing interests.

Footnotes

Publisher’s note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

References

  • 1.Asherman JG. Amenorrhoea traumatica (atretica) BJOG. 1948;55(1):23–30. doi: 10.1111/j.1471-0528.1948.tb07045.x. [DOI] [PubMed] [Google Scholar]
  • 2.Asherman JG. Traumatic intrauterine adhesions. BJOG. 1950;57(6):892–896. doi: 10.1111/j.1471-0528.1950.tb06053.x. [DOI] [PubMed] [Google Scholar]
  • 3.Yu D, Wong YM, Cheong Y, Xia E, Li TC. Asherman syndrome—one century later. Fertil Steril. 2008;89:759–779. doi: 10.1016/j.fertnstert.2008.02.096. [DOI] [PubMed] [Google Scholar]
  • 4.Senturk LM, Erel CT. Thin endometrium in assisted reproductive technology. Curr Opin Obstet Gynecol. 2008;20:221–228. doi: 10.1097/GCO.0b013e328302143c. [DOI] [PubMed] [Google Scholar]
  • 5.Mouhayar Y, Sharara F. Modern management of thin lining. Middle East Fertil Soc J. 2017;22:1–2. doi: 10.1016/j.mefs.2016.09.001. [DOI] [Google Scholar]
  • 6.Gargett CE, Ye L. Endometrial reconstruction from stem cells. Fertil Steril. 2012;98:1120. doi: 10.1016/j.fertnstert.2012.05.004. [DOI] [PubMed] [Google Scholar]
  • 7.Ratajczak MZ, Zuba-Surma EK, Machalinski B, Ratajczak J, Kucia M. Very small embryonic-like (VSEL) stem cells: purification from adult organs, characterization, and biological significance. Stem Cell Rev. 2008;4:89–99. doi: 10.1007/s12015-008-9018-0. [DOI] [PubMed] [Google Scholar]
  • 8.Franz RW, Parks A, Shah KJ, Hankins T, Hartman JF, Wright ML. Use of autologous bone marrow mononuclear cell implantation therapy as a limb salvage procedure in patients with severe peripheral arterial disease. J Vasc Surg. 2009;50:1378–1390. doi: 10.1016/j.jvs.2009.07.113. [DOI] [PubMed] [Google Scholar]
  • 9.Yanishi K, Shoji K, Fujioka A, Hori Y, Yukawa A, Matoba S. Impact of therapeutic angiogenesis using autologous bone marrow-derived mononuclear cell implantation in patients with no-option critical limb ischemia. Ann Vasc Dis. 2020;13:13–22. doi: 10.3400/avd.ra.20-00002. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 10.Beltran-Camacho L, Jimenez-Palomares M, Torres-Rojas M, Perez-Segura MC, Serrano A, Sanchez-Gomar I, Antonio Rosal-Vela A, Eslava-Alcon S, Perez-Segura MC, Serrano A, Antequera-González B, Alonso-Piñero JA, González-Rovira A, Extremera-García MJ, Rodriguez-Piñero M, Moreno-Luna R, Larsen MR, Larsen MR, Durán-Ruiz MC. Identification of the initial molecular changes detected in response to circulating angiogenic cellsmediated therapy in critical limb ischemia. Stem Cell Res Ther. 2020;11:106. doi: 10.1186/s13287-020-01591-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 11.Nossent AY, Bastiaansen AJ, Peters EA, de Vries MR, Aref Z, Welten SM, de Jager SCA, van der Pouw KTCTM, Quax PHA. CCR7-CCL19/CCL21 axis is essential for effective arteriogenesis in a murine model of hindlimb ischemia. J Am Heart Assoc. 2017;6:e005281. doi: 10.1161/JAHA.116.005281. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 12.Lin RZ, Lee CN, Moreno-Luna R, Neumeyer J, Piekarski B, Zhou P, Moses MA, Sachdev M, Pu WT, Emani S, Melero-Martin JM. Host non-inflammatory neutrophils mediate the engraftment of bioengineered vascular networks. Nat Biomed Eng. 2017;1:0081. doi: 10.1038/s41551-017-0081. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 13.Seignez C, Phillipson M. The multitasking neutrophils and their involvement in angiogenesis. Curr Opin Hematol. 2017;24:3–8. doi: 10.1097/MOH.0000000000000300. [DOI] [PubMed] [Google Scholar]
  • 14.Korbling M, Estrov Z. Adult stem cells for tissue repair—a new therapeutic concept? N Engl J Med. 2003;349:570–582. doi: 10.1056/NEJMra022361. [DOI] [PubMed] [Google Scholar]
  • 15.Du HL, Taylor HS. Stem cells and reproduction. Curr Opin Obstet Gynecol. 2010;22:235–241. doi: 10.1097/GCO.0b013e328338c152. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 16.Du H, Taylor HS. Contribution of bone marrow-derived stem cells to endometrium and endometriosis. Stem Cells. 2007;25:2082–2086. doi: 10.1634/stemcells.2006-0828. [DOI] [PubMed] [Google Scholar]
  • 17.Cervello I, Martinez-Conejero JA, Horcajadas JA, Pellicer A, Simo’n C. Identification, characterization and co-localization of label-retaining cell population in mouse endometrium with typical undifferentiated markers. Hum Reprod. 2007;22:45–51. doi: 10.1093/humrep/del332. [DOI] [PubMed] [Google Scholar]
  • 18.Mints M, Jansson M, Sadeghi B, Westgren M, Uzunel M, Hassan M, Palmblad J. Endometrial endothelial cells are derived from donor stem cells in a bone marrow transplant recipient. Hum Reprod. 143. 10.1093/humrep/dem342. [DOI] [PubMed]
  • 19.Du H, Taylor HS. Stem cells and female reproduction. Reprod Sci. 2009;16:126–139. doi: 10.1177/1933719108329956. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 20.Du H, Naqvi H, Taylor HS. Ischemia/reperfusion injury promotes and granulocytecolony stimulating factor inhibits migration of bone marrow-derived stem cells to endometrium. Stem Cells Dev. 2012;21:3324–3331. doi: 10.1089/scd.2011.0193. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 21.Morelli SS, Rameshwar P, Goldsmith LT. Experimental evidence for bone marrow as a source of nonhematopoietic endometrial stromal and epithelial compartment cells in a murine model. Biol Reprod. 2013;89:7. doi: 10.1095/biolreprod.113.107987. [DOI] [PubMed] [Google Scholar]
  • 22.Alawadhi F, Du H, Cakmak H, Taylor HS. Bone marrow-derived stem cell (BMDSC) transplantation improves fertility in a murine model of Asherman’s syndrome. PLoS One. 2014;9:e96662. doi: 10.1371/journal.pone.0096662. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 23.Kilic S, Yuksel B, Pinarli F, Albayrak A, Boztok B, DelibasiT. Effect of stem cell application on Asherman syndrome, an experimental rat model. J Assist Reprod Genet. 2014;31:975–982. doi: 10.1007/s10815-014-0268-2. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 24.Taylor HS. Endometrial cells derived fromdonor stem cells in bone marrow transplant recipients. JAMA. 2004;292:81–85. doi: 10.1001/jama.292.1.81. [DOI] [PubMed] [Google Scholar]
  • 25.Nagori CB, Panchal SY, Patel H. Endometrial regeneration using autologous adult stem cells followed by conception by in vitro fertilization in a patient of severe Asherman’s syndrome. J Hum Reprod Sci. 2011;4:43–48. doi: 10.4103/0974-1208.82360. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 26.Gargett CE, Healy DL. Generating receptive endometrium in Asherman's syndrome. J Hum Reprod Sci. 2011;4:49–52. [PMC free article] [PubMed] [Google Scholar]
  • 27.Santamaria X, Cabanillas S, Cervello I, Arbona C, Raga F, Ferro J, Palmero J, Remohi J, Pellicer A, Simon C. Autologous cell therapy with CD133 bonemarrow-derived stem cells for refractory Asherman’s syndrome andendometrial atrophy: a pilot cohort study. Hum Reprod. 2016;31(5):1087–1096. doi: 10.1093/humrep/dew042. [DOI] [PubMed] [Google Scholar]
  • 28.Singh N, Mohanty S, Seth T, Shankar M, Bhaskaran S, Dharmendra S. Autologous stem cell transplantation in refractory Asherman’s syndrome: a novel cell-based therapy. J Hum Reprod Sci. 2014;7:93–98. doi: 10.4103/0974-1208.138864. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 29.Singh N, Shekhar B, Mohanty S, Kumar S, Seth T, Girish B. Autologous bone marrow derived stem cell therapy for Asherman's syndrome and endometrial atrophy: a 5-year follow-up study. J Hum Reprod Sci. 2020;13:31–37. doi: 10.4103/jhrs.JHRS_64_19. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 30.Benor A, Gay A, De Cherney A. An update on stem cell therapy for Asherman syndrome. J Asist Reprod Genet. 2020;37:1511–1529. doi: 10.1007/s10815-020-01801-x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 31.Gharibeh N, Aghebati-Maleki L, Madani J, Pourakbari R, Yousefi M, Javad AH. Cell-based therapy in thin endometrium and Asherman syndrome. Stem Cell Res Ther. 2022;13:33. doi: 10.1186/s13287-021-02698-832. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 32.March C, Israel R, March A. Hysteroscopic management of intrauterine adhesions. Am J Obstet Gynecol. 1978;11:653–657. doi: 10.1016/0002-9378(78)90322-8. [DOI] [PubMed] [Google Scholar]
  • 33.Prockop DJ. Repair of tissues by adult stem/progenitor cells (MSCs): controversies, myths, and changing paradigms. Mol Ther. 2009;17:939–946. doi: 10.1186/s13287-02102698-8. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 34.Caplan AI. Why are MSCs therapeutic? New data: new insight. J Pathol. 2009;217:31–24. doi: 10.1002/path.2469. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 35.Fadini GP, Losordo D, Dimmeler S. Critical reevaluation of endothelial progenitor cell phenotypes for therapeutic and diagnostic use. Circ Res. 2012;110(4):624–637. doi: 10.1161/CIRCRESAHA.111.243386. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 36.Wei W, Li L, Lin D, Zhong-Jing W, Jing-Jian D, Xiao-Yu L, Ting J, Li W, Hong-Xiang W, Hong M, Shi Z. Autologous bone marrow mononuclear cell transplantation therapy ımproved symptoms in patients with refractory diabetic sensorimotor polyneuropathy via the mechanisms of paracrine and ımmunomodulation: a controlled study. Cell Transplantation. 2020;29:1–11. doi: 10.1177/0963689720949258. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 37.Kwee BJ, Budina E, Najibi AJ, Mooney DJ. CD4 T-cells regulate angiogenesis and myogenesis. Biomaterials. 2018;178:109–121. doi: 10.1016/j.biomaterials.2018.06.003. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 38.Dehkordi AN, Mirahmadi Babaheydari FM, Chehelgerdi M, Dehkordi SR. Skin tissue engineering: wound healing based on stem-cell-based therapeutic strategies. Stem Cell Res Ther. 2019;29(1):111. doi: 10.1186/s13287-019-1212-2. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 39.Barash A, Dekel N, Fieldust S, Segal I, Schechtman E, Granot I. Local injury to the endometrium doubles the incidence of successful pregnancies in patients undergoing in vitro fertilization. Fertil Steril. 2003;79:1317–1322. doi: 10.1016/s0015-0282(03)00345-5. [DOI] [PubMed] [Google Scholar]
  • 40.Zhou L, Li R, Wang R, Huang H, Zhong BA. Local injury to the endometrium in controlled ovarian hyperstimulation cycles improves implantation rates. Fertil Steril. 2008;89:1166–1176. doi: 10.1016/j.fertnstert.2007.05.064. [DOI] [PubMed] [Google Scholar]
  • 41.Wysoczynki M, Khan A, Bolli B. New paradigms in cell therapy: repeated dosing, intravenous delivery, immunomodulatory actions, and new cell types. Circ Res. 2018;123(2):138–158. doi: 10.1161/CIRCRESAHA.118.313251. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 42.Bolli R. Repeated cell therapy: a paradigm shift whose time has come. Circ Res. 2017;120:1072–1074. doi: 10.1161/CIRCRESAHA.117.310710. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 43.Tokita Y, Tang XL, Li Q, Wysoczynski M, Hong KU, Nakamura S, Wu WJ, Xie W, Li D, Hunt G, Ou Q, Stowers H, Bolli R. Repeated administrations of cardiac progenitor cells are markedly more effective than a single administration: a new paradigm in cell therapy. Circ Res. 2016;119:635–651. doi: 10.1161/CIRCRESAHA.116.308937. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 44.Guo Y, Wysoczynski M, Nong Y, Tomlin A, Zhu X, Gumpert AM, Nasr M, Muthusamy S, Li H, Book M, Khan A, Hong KU, Li Q, Bolli R. Repeated doses of cardiac mesenchymal cells are therapeutically superior to a single dose in mice with old myocardial infarction. Basic Res Cardiol. 2017;112:18. doi: 10.1007/s00395-017-0606-5. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 45.Jarocha D, Zuba-Surma E, Majka M. Dimethyl sulfoxide (DMSO) increases percentage of CXCR4(+) hematopoietic stem/progenitor cells, their responsiveness to an SDF-1 gradient, homing capacities, and survival. Cell Transplant. 2016;25(7):1247–1257. doi: 10.3727/096368915X689424. [DOI] [PubMed] [Google Scholar]
  • 46.Huang J, Li Q, Yuan X, Liu Q, Zhang W, Li P. Intrauterine infusion of clinically graded human umbilical cord-derived mesenchymal stem cells for the treatment of poor healing after uterine injury: a phase I clinical trial. Stem Cell Res Ther. 2022;13:85. doi: 10.1186/s13287-022-02756-9. [DOI] [PMC free article] [PubMed] [Google Scholar]

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